This project studies peripheral blood progenitors (PBP) as a target for retrovirus mediated gene therapy of inherited diseases affecting the function of human phagocytic cells, including neutrophils, eosinophils, and monocytes. Studies of growth factors which influence the differentiation of PBPs are also a part of this project. PBPs which may include totipotent stem cells are similar or identical to the blood precursors in bone marrow, but circulate in small numbers in peripheral blood. PBP harvested and concentrated from peripheral blood by centrifugation apheresis have been used by others as an autologous transplant in lieu of or combined with progenitors harvested from bone marrow to reconstitute the bone marrow of patients with cancer treated with cytotoxic chemotherapy. PBP harvested from patients with inherited immune diseases such as chronic granulomatous disease or other immune deficiency are a readily available target for functional correction by gene transfer. We purified PBP from apheresis derived leukocytes and studied conditions for differentiating them into neutrophils, monocytes, and/or eosinophils using both liquid and soft agarose cultures. In one series of studies we demonstrated that stem cell factor (also called c- kit ligand) was not active alone, but when combined with other hematopoietic factors greatly increased both the number and size of myeloid colonies derived from PBP. In related studies we demonstrated that the eosinophil factor IL5 alone would not support colony growth of PBPs in the early stages of differentiation, but that IL5 alone would support colony growth after culturing PBPs for 4 days in early acting growth factors. This suggests that responsiveness to late acting hematopoietic factors may require the expression of receptors to such factors. This type of information is essential to the goal of proliferation of hematopoietic progenitors in vitro for purposes of gene therapy or transplantation. Studies of the number of circulating PBP indicated that this is a highly regulated and constant parameter, but that endotoxin administration, a model of gram negative bacterial infection, increased the number of circulating PBP five-fold. In other studies using the retroviral constructs we determined optimum conditions for transfer of genes into PBP. Determination of such conditions are essential to the development of protocols for gene therapy for immune system disorders affecting bone marrow.